The basal keratinocytes of all stratified squamous epithelia appear to have a common mechanism that triggers irreversible loss of proliferative potential ("commitment") at a time closely linked to migration upward into the suprabasal compartment where they undergo one of several alternate patterns of differentiation. These distinctive forms of differentiation (e.g.,keratinization vs. nonkeratinization) expressed by epidermal vs. oral mucosal keratinocytes are the result of keratinocyte subtype-specific mechanisms that specify expression of alternate sets of differentiation- related genes, such as keratins, filaggrin, and loricrin. Our recent studies have demonstrated that, despite the retinoid sensitivity of epidermal differentiation and the differential expression of one RAR isoform, RARbeta, by the various types of stratified squamous epithelia, neither retinoids nor RARbeta plays a determining role in the mechanism that specifies the choice of alternate pathways of suprabasal differentiation. We propose to determine the roles in keratinocyte differentiation of a potential keratinization pathway-specific transcriptional regulator PPAR-alpha, and of potential common "commitment"-triggering transcriptional regulators of the AP1 family -- c- fos, c-jun, and the EB virus-encoded, AP1-related protein Zta. We will use retroviral vectors to generate stable transductants of normal human keratinocytes and squamous cell carcinoma cells that overexpress these proteins, either constitutively or under control of a tetracycline- regulated promoter. We will characterize the transductants for alterations in proliferation, keratin expression, and histogenic potential in conventional and organotypic culture. This study will test several important hypotheses: 1) that the recently characterized nuclear hormone receptor PPAR-alpha, which is activated by lipid ligands, modulates expression of suprabasal differentiation proteins, 2) that Zta induces growth arrest and expression of differentiation by keratinocytes, thereby providing a permissive environment for the lytic, productive phase of Herpes infection, and 3) that expression of c-fos or c-jun triggers irreversible growth arrest and commitment to differentiation in keratinocytes. This pilot and feasibility study will also permit us to establish expertise in the use of the recently developed retroviral-based tet-regulated gene expression system and will help us identify novel keratinocyte differentiation regulatory system on which to base more extensive studies in the future.